OBTAINING AND CHARACTERIZATION OF RECOMBINANT FLUORESCENT DERIVATIVES OF SOLUBLE HUMAN HB-EGF

Abstract

Heparin-binding EGF-like growth factor (HB-EGF) belongs to the epidermal growth factor receptor family and is synthesized as a transmembrane precursor proHB-EGF. Binding of sHB-EGF with receptors EGFR and HER-4 results in formation of ligand-receptor complexes and activation of signaling pathways and thereby promotes survival, proliferation and migration of cells. The aim of the study was to obtain the recombinant fluorescent derivatives of full-length soluble sHB-EGF – mCherry-sHB-EGF and truncated sHB-EGFΔ84-106 (without heparin-binding domain) – mCherry-sHB-EGFΔ84-106. The recombinant fluorescent derivatives may be used to investigate the sHB-EGF binding with receptors EGFR and HER-4, intracellular transportation of ligand-receptor complexes and the role of sHB-EGF heparin-binding domain in sHB-EGF biological activity. It was shown the ability of both fluorescent derivatives specifically bind to EGFR, to internalize in receptor-mediated pathway and to enhance the proliferation of 3T3 cells. Absence of heparin-binding domain in structure of mCherry-sHB-EGFΔ84-106 significantly did not effect on its ability to bind with receptor, but decreased twice its mitogenic activity. Thus, obtained fluorescent derivatives of sHB-EGF could be a convenient tool for investigation of molecular mechanisms of sHB-EGF biological activity and role of heparin-binding domain in these processes.

Keywords

• heparin-binding EGF-like growth factor (HB-EGF)
• epidermal growth factor receptor.